Heme- and iron-scavenging systems are important virulence factors among a diversity of pathogenic bacteria. The current proposal, however, will examine bacterial heme acquisition from an ecological point of view, according to the established premise that competition for limiting nutrients is a fundamental factor influencing microbial ecosystems. Gingival crevicular fluid (GCF) is essentially a plasma transudate and, because plasma normally contains very low concentrations of heme and several specific heme-binding proteins, this growth factor may be of limited availability to bacteria inhabiting the healthy gingival crevice. The heme-requiring anaerobe Prevotella intermedia may be isolated from relatively healthy gingivae (albeit in low relative numbers) whereas the more pathogenic Porphyromonas gingivalis is rarely found in either healthy or mildly inflamed gingivae but is highly associated with some forms of adult periodontitis. The distribution of these two organisms may reflect their relative heme-sequestering capacities, suggesting that P.intermedius competitively suppresses the growth of P.gingivalis under conditions of relative periodontal health (heme restriction). The current proposal will develop a radiolabeled hemin-uptake assay with which to compare heme-acquisition by P.intermedia and P.gingivalis. The assay will be modified to examine the influence of the specific heme- binding proteins, human serum albumin and human hemopexin, on bacterial heme-acquisition. The high affinity with which these proteins bind their ligands may restrict heme-uptake by bacteria. Preliminary studies aimed at determining the mechanisms of bacterial heme-acquisition, such as the nature of the receptors, will be undertaken. Investigations will also be aimed at developing a plasma-based culture system with which to examine nutritional and other interactions among periodontal bacteria. Plasma will be used as the closest available approximation to GCF. Identification of restrictive factors (either bactericidal or nutritional) preventing growth of specific bacteria may relate directly to periodontal ecology. Similarly, identification of organisms able to overcome the restrictive factors and establish support for other, potentially pathogenic strains, will be an important step in elucidating the ecological progression of gingival inflammatory conditions. Evidence suggesting that heme availability and host heme-binding proteins influence subgingival microbial ecology will justify future studies aimed at: (i) correlating the plasma and GCF concentrations of host heme-binding proteins with the proportions of heme-requiring anaerobes and periodontal status of individuals; (ii) ascertaining the mechanisms and regulation of bacterial heme-acquisition; (iii) understanding the ecological factors favoring ascendancy of presumed periodontal pathogens using a plasma-based continuous culture model.